Monitored changes in the number of copies of a gene during DNA replication control the timing of sporulation in bacteria. This discovery links replication to the concept that a gene's location on a chromosome can influence cell traits.
For decades it has been known1 that the location of a gene on its chromosome can influence the level at which it is expressed. Most bacterial chromosomes are circular, and their replication begins at a single bidirectional origin. As such, during chromosome replication, genes close to the origin of replication will be transiently present in more copies (present at a higher dosage) than those close to the terminus of replication. Altering the distance of a gene from the origin of replication systematically alters its level of expression during the cell's replication cycle2,3,4. But until now, the significance of gene location has largely focused on whether highly expressed genes are preferentially located in the origin-proximal half of the chromosome, because this provides the cell with a growth advantage due to a positive gene-dosage effect5. Writing in Cell, Narula et al.6 report a new twist on the role of chromosomal location in gene function, in coordinating sporulation with chromosome replication in the bacterium Bacillus subtilis.
When starved, B. subtilis can initiate a cascade of protein phosphorylation that leads to sporulation, producing a dormant spore that is resistant to starvation conditions and that can eventually resume growth under favourable conditions. The first components of this phosphorelay are a kinase enzyme called KinA and a response-regulator protein, Spo0F. In vitro evidence7 has suggested that, although phosphorylation of Spo0F by KinA is necessary for the activation of early sporulation genes, high concentrations of Spo0F can also inhibit the activity of KinA. Narula et al. confirmed this result in vivo, demonstrating that high levels of Spo0F induce a negative-feedback loop that inhibits the phosphorelay.
The spo0F gene is located near the origin of replication, whereas the kinA gene is located close to the replication terminus. Narula et al. report that the positions of spo0F and kinA seem to be crucial for their ability to efficiently regulate sporulation. Because of their respective locations, during replication there is a temporary twofold increase in the dosage of spo0F relative to kinA (Fig. 1a). By using computer simulations and then verifying their models in vivo, the authors showed that the transient increase in Spo0F concentration inhibits KinA until replication is completed, leading to pulsing dynamics of sporulation-gene expression during each cell cycle (Fig. 1b). Cells will only sporulate once they cross a threshold level of sporulation-gene expression, which is achieved through a positive-feedback loop that increases levels of KinA concentration — a process that takes several rounds of cell division8.
Narula and colleagues then performed translocation experiments, in which they moved spo0F or kinA towards the terminus or origin of replication, respectively. These translocations abolished pulsing, confirming that a transient imbalance in the dosage of the two genes is necessary for pulsing of early sporulation-gene expression and for proper coordination of the sporulation program with DNA replication. These data, together with the authors' finding that the relative locations of kinA and spo0F are similar in 45 other species of sporulating bacteria, show for the first time that the siting of interacting genes at different locations on the chromosome could have evolved as a way of controlling how the gene products function.
Monitoring chromosome replication status is crucial for many species. In the case of B. subtilis, initiation of sporulation without complete chromosomes for both the mother cell and the future spore cell would be a waste of resources. It has long been known9 that a checkpoint is activated to inhibit sporulation when DNA is damaged or replication is defective. Narula and colleagues have identified a remarkably simple mechanism by which cells can monitor the replication status of the chromosome.
The regulatory mechanism presented in this study deepens our understanding of the potential variety of mechanisms that might regulate changes in cellular traits. But the work also raises several interesting avenues for further investigation. For example, it is unclear whether this particular situation is a biological one-off. It seems more likely that there are other traits, both in B. subtilis and in other organisms, that are regulated by temporal variations in gene-product ratios associated with gene location.
It also remains to be seen whether more-complex versions of this mechanism exist, involving more than two genes, and whether such mechanisms could be involved in replication fidelity. For instance, could this type of regulatory mechanism act as a brake on chromosomal rearrangements such as inversions, which might disrupt the relative locations of genes in regulatory networks that rely on dosage imbalances?
Narula and colleagues' work illustrates the potential importance of gene location in perhaps unexpected aspects of cell biology. It will doubtless motivate future experiments in chromosome remodelling. Perhaps it will also prompt a re-examination of old data, to assess whether arbitrary choices made in genetic engineering might have affected experimental outcomes.
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